Klenow fragment

Klenow fragment

The Klenow fragment is a large protein fragment produced when DNA polymerase I from "E. coli" is enzymatically cleaved by the protease subtilisin. First reported in 1970 [cite journal| author=Klenow H and Henningsen I | title=Selective Elimination of the Exonuclease Activity of the Deoxyribonucleic Acid Polymerase from Escherichia coli B by Limited Proteolysis| journal= Proc Natl Acad Sci| volume= 65 |pages=168–175 | year=1970 | pmid=4905667 | doi = 10.1073/pnas.65.1.168 | url=http://www.pubmedcentral.nih.gov/articlerender.fcgi?tool=pubmed&pubmedid=4905667] , it retains the 5’ → 3’ polymerase activity and the 3’ → 5’ exonuclease activity for removal of precoding nucleotides and proofreading, but loses its 5' → 3' exonuclease activity.

The other smaller fragment formed when DNA polymerase I from "E. coli" is cleaved by subtilisin retains the 5'→ 3' exonuclease activity but does not have the other two activities exhibited by the Klenow fragment (i.e. 5'-> 3' polymerase activity, and 3'->5' nuclease activity).

Research

Because the 5' → 3' exonuclease activity of DNA polymerase I from "E. coli" makes it unsuitable for many applications, the Klenow fragment, which lacks this activity, can be very useful in research. The Klenow fragment is extremely useful for research-based tasks such as:

* Synthesis of double-stranded DNA from single-stranded templates
* Filling in recessed 3' ends of DNA fragments
* Digesting away protruding 3' overhangs
* Preparation of radioactive DNA probes

The Klenow fragment was also the original enzyme used for greatly amplifying segments of DNA in the polymerase chain reaction (PCR) processcite journal | author=Saiki RK et al. | title=Enzymatic Amplification of β-globin Genomic Sequences and Restriction Site Analysis for Diagnosis of Sickle Cell Anemia| journal= Science| volume= 230| pages=1350-54 | year= 1985 | pmid = 2999980 | doi = 10.1126/science.2999980 ] , before being replaced by thermostable enzymes such as Taq polymerasecite journal | author=Saiki et al. | title=Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase | journal= Science | volume= 239| pages=487-91 | year=1988 | pmid = 2448875 | doi = 10.1126/science.2448875 ] .

The exo- Klenow fragment

Just as the 5' → 3' exonuclease activity of DNA polymerase I from "E.coli" can be undesirable, the 3' → 5' exonuclease activity of Klenow fragment can also be undesirable for certain applications. This problem can be overcome by introducing mutations in the gene that encodes Klenow. This results in forms of the enzyme being expressed that retain 5' → 3' polymerase activity, but lack any exonuclease activity (5' → 3' or 3' → 5'). This form of the enzyme is called the exo- Klenow fragment.

References

External links

*
* [http://www.vivo.colostate.edu/hbooks/genetics/biotech/enzymes/klenow.html Diagram at vivo.colostate.edu]


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Look at other dictionaries:

  • Klenow fragment — Klenow fragment. См. фрагмент Кленова. (Источник: «Англо русский толковый словарь генетических терминов». Арефьев В.А., Лисовенко Л.А., Москва: Изд во ВНИРО, 1995 г.) …   Молекулярная биология и генетика. Толковый словарь.

  • Klenow-Fragment — Als Klenow Fragment, auch Klenow Enzym, wird das größere der beiden Proteinfragmente der DNA Polymerase I aus Escherichia coli bezeichnet, die nach enzymatischer Spaltung mit Subtilisin entstehen. Es besitzt noch die 5 →3 Polymerase Aktivität und …   Deutsch Wikipedia

  • Klenow fragment — Larger part of the bacterial DNA polymerase I (76 kD) that remains after treatment with subtilisin; retains some but not all exonuclease and polymerase activity …   Dictionary of molecular biology

  • Klenow fragment — A truncated form of DNA polymerase I from E. coli, used extensively for the production of synthetic DNA molecules as it retains polymerase and 3 exonuclease activities, but not 5 exonuclease activity …   Glossary of Biotechnology

  • Klenow-Enzym — Als Klenow Fragment bzw. Klenow Enzym wird die größere der beiden Untereinheiten der DNA Polymerase I von Escherichia coli bezeichnet. Es wurde nach seinem Entdecker, dem dänischen Biochemiker Hans Klenow benannt. Eigenschaften Es verfügt über… …   Deutsch Wikipedia

  • Klenow-Polymerase — Als Klenow Fragment bzw. Klenow Enzym wird die größere der beiden Untereinheiten der DNA Polymerase I von Escherichia coli bezeichnet. Es wurde nach seinem Entdecker, dem dänischen Biochemiker Hans Klenow benannt. Eigenschaften Es verfügt über… …   Deutsch Wikipedia

  • Fragment de Klenow — Le fragment de Klenow est le plus gros des 2 fragments protéiques formés par l hydrolyse de l ADN polymérase I d Escherichia coli par une protéase (subtilisine). Découvert en 1970, il conserve deux des activités catalytiques de l ADN polymérase… …   Wikipédia en Français

  • fragment — A small part broken from a larger entity. acentric f. SYN: acentric chromosome. Brimacombe f. a ribonucleoprotein f. obtained by mild ribonuclease treatment of ribosomes. butterfly f. a broad triangular f. that is commonly present in …   Medical dictionary

  • Kf — Klenow fragment …   Medical dictionary

  • Kf — • Klenow fragment …   Dictionary of medical acronyms & abbreviations

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